Polycystic ovary syndrome (PCOS) is the most common endocrine disorder in women of reproductive age. It is associated with profound insulin resistance resulting in a markedly increased risk for non-insulin dependent diabetes mellitus (NIDDM) at a strikingly early age (3rd-4th decades). The overall hypothesis of this research is that insulin resistance in PCOS is the result of several distinctive defects in insulin receptor-mediated signaling. Further, we propose that each defect has a unique genetic basis compared to defects producing insulin resistance associated with typical NIDDM, with obesity, or with the rare syndromes of extreme insulin resistance. We have found a novel abnormality of insulin- receptor signaling (increased insulin-independent insulin receptor serine phosphorylation and decreased insulin-stimulated insulin receptor tyrosine phosphorylation) in association with insulin resistance in approximately 50% of PCOS women. This defect is present in the major insulin target tissue, muscle, and persists in cultured cells suggesting a genetic abnormality. The Specific Aims of this proposal are, thus: 1) To determine the role of insulin receptor serine phosphorylation in the pathogenesis of insulin resistance in PCOS. We hypothesize that increased insulin- independent insulin receptor serine phosphorylation inhibits insulin- induced receptor-mediated signaling in PCOS. This will be investigated by examining the effects of dephosphorylation on the kinase activity of insulin receptors partially-purified from muscle and from fat, using serine specific (e.g. phosphatase type-2A) as well as nonspecific (e.g. alkaline phosphatase) phosphatases. 2) To determine whether there are post-insulin receptor signaling defects in PCOS. In PCOS there is a marked shift to the right in the insulin dose-response curve for glucose uptake in fat and in muscle as well as a decrease in adipocyte GLUT4 content. However, approximately 30% of such insulin resistant PCOS women have normal insulin receptor phosphorylation, and we hypothesize that these women have downstream signaling defects. Moreover, we hypothesize that 50% of PCOS women with defects in insulin receptor phosphorylation will also have decreased insulin-receptor mediated downstream signaling. This will be investigated by assessing insulin stimulation, in vivo and in intact adipocytes, of phosphatidylinositol 3-kinase (PtdIns 3-kinase) activity. Glucose uptake and GLUT4 content will also be determined. The ED50 insulin and Vmax for glucose use and for PtdIns 3-kinase activation will be examined. 3) To determine whether defects in insulin action in PCOS are genetic. If insulin resistance in PCOS is a genetic defect, first degree relatives should be affected. This will be investigated by determining total body and cellular insulin action in brothers of PCOS probands. We will screen for mutations in the insulin receptor and IRS-1 genes of PCOS women with denaturing gradient gel electrophoresis or single stranded conformation polymorphisms. Lean and obese PCOS women, brothers of PCOS women and age-, weight-, ethnicity- and sex-matched normal control subjects will participate in these studies.